SummaryEach year millions of animals undertake remarkable migratory journeys, across oceans and through hemispheres, guided by the Earth’s magnetic field. The cellular and molecular basis of this enigmatic sense, known as magnetoreception, remains an unsolved scientific mystery. One hypothesis that attempts to explain the basis of this sensory faculty is known as the magnetite theory of magnetoreception. It argues that magnetic information is transduced into a neuronal impulse by employing the iron oxide magnetite (Fe3O4). Current evidence indicates that pigeons employ a magnetoreceptor that is associated with the ophthalmic branch of the trigeminal nerve and the vestibular system, but the sensory cells remain undiscovered. The goal of this ambitious proposal is to discover the cells and molecules that mediate magnetoreception. This overall objective can be divided into three specific aims: (1) the identification of putative magnetoreceptive cells (PMCs); (2) the cellular characterisation of PMCs; and (3) the discovery and functional ablation of molecules specific to PMCs. In tackling these three aims this proposal adopts a reductionist mindset, employing and developing the latest imaging, subcellular, and molecular technologies.

Each year millions of animals undertake remarkable migratory journeys, across oceans and through hemispheres, guided by the Earth’s magnetic field. The cellular and molecular basis of this enigmatic sense, known as magnetoreception, remains an unsolved scientific mystery. One hypothesis that attempts to explain the basis of this sensory faculty is known as the magnetite theory of magnetoreception. It argues that magnetic information is transduced into a neuronal impulse by employing the iron oxide magnetite (Fe3O4). Current evidence indicates that pigeons employ a magnetoreceptor that is associated with the ophthalmic branch of the trigeminal nerve and the vestibular system, but the sensory cells remain undiscovered. The goal of this ambitious proposal is to discover the cells and molecules that mediate magnetoreception. This overall objective can be divided into three specific aims: (1) the identification of putative magnetoreceptive cells (PMCs); (2) the cellular characterisation of PMCs; and (3) the discovery and functional ablation of molecules specific to PMCs. In tackling these three aims this proposal adopts a reductionist mindset, employing and developing the latest imaging, subcellular, and molecular technologies.

SummaryRecent advances in genome sequencing illustrate the complexity, heterogeneity and plasticity of cancer genomes. In leukemia - a group of blood cancers affecting 300,000 new patients every year – we know over 100 driver mutations. This genetic complexity poses a daunting challenge for the development of targeted therapies and highlights the urgent need for evaluating them in combination. One gene class that has recently emerged as highly promising target space are chromatin regulators, which maintain aberrant cell fate programs in leukemia. The dependency on altered chromatin states is thought to provide great therapeutic opportunities, since epigenetic aberrations are reversible and controlled by a machinery that is amenable to drug modulation. However, the precise mechanisms underlying these dependencies and the most effective and safe targets to exploit them therapeutically remain unknown.
Here we propose an innovative approach combining genetically engineered leukemia mouse models and advanced in-vivo RNAi technologies to explore chromatin-associated vulnerabilities at an unprecedented level of depth. Following a first screen in MLL-AF9;Nras-driven AML, which led to the discovery of BRD4 as a promising therapeutic target, we aim to (1) construct a knockdown-validated shRNA library targeting 520 chromatin regulators and use it to comparatively probe chromatin-associated dependencies in diverse leukemia subtypes; (2) explore the mechanistic basis of response and resistance to suppression of BRD4 and new chromatin-associated targets; and (3) pioneer a system for multiplexed combinatorial RNAi screening and use it to identify synergies between established and new chromatin-associated targets. We envision that this ERC-funded project will generate a comprehensive functional-genetic dataset that will greatly complement ongoing genome and epigenome profiling studies and ultimately guide the development of targeted therapies for leukemia and, potentially, other cancers.

Recent advances in genome sequencing illustrate the complexity, heterogeneity and plasticity of cancer genomes. In leukemia - a group of blood cancers affecting 300,000 new patients every year – we know over 100 driver mutations. This genetic complexity poses a daunting challenge for the development of targeted therapies and highlights the urgent need for evaluating them in combination. One gene class that has recently emerged as highly promising target space are chromatin regulators, which maintain aberrant cell fate programs in leukemia. The dependency on altered chromatin states is thought to provide great therapeutic opportunities, since epigenetic aberrations are reversible and controlled by a machinery that is amenable to drug modulation. However, the precise mechanisms underlying these dependencies and the most effective and safe targets to exploit them therapeutically remain unknown.
Here we propose an innovative approach combining genetically engineered leukemia mouse models and advanced in-vivo RNAi technologies to explore chromatin-associated vulnerabilities at an unprecedented level of depth. Following a first screen in MLL-AF9;Nras-driven AML, which led to the discovery of BRD4 as a promising therapeutic target, we aim to (1) construct a knockdown-validated shRNA library targeting 520 chromatin regulators and use it to comparatively probe chromatin-associated dependencies in diverse leukemia subtypes; (2) explore the mechanistic basis of response and resistance to suppression of BRD4 and new chromatin-associated targets; and (3) pioneer a system for multiplexed combinatorial RNAi screening and use it to identify synergies between established and new chromatin-associated targets. We envision that this ERC-funded project will generate a comprehensive functional-genetic dataset that will greatly complement ongoing genome and epigenome profiling studies and ultimately guide the development of targeted therapies for leukemia and, potentially, other cancers.

SummaryAn animal’s decision on how to respond to the environment is based not only on the sensory information available, but further depends on internal factors such as stress, sleep / wakefulness, hunger / satiety and experience. Neurotransmitters and neuropeptides in the brain modulate neural circuits accordingly so that appropriate behaviors are generated. Aberrant neuromodulation is implicated in diseases such as insomnia, obesity or anorexia. Given the complexity of most neural systems studied, we lack good models of how neuromodulators systemically affect the activities of neural networks.
To overcome this problem, I propose to study neural circuits in the nematode C. elegans, which is a genetically tractable model organism with a simple and anatomically defined nervous system. I will focus on the neural circuits involved in oxygen chemosensory behaviors. Worms can smell oxygen and they use this information to navigate through heterogeneous environments. This enables them to find food and to engage in social interactions. Oxygen chemosensory behaviors are highly modulated by experience and nutritional status, but the underlying mechanisms are not understood.
I established behavioral assays that allow studying the modulation of oxygen behaviors in a rigorously quantifiable manner. I also acquired expertise in micro-fabrication technologies and developed imaging devices to measure the activity of neurons in live animals. The first two aims of this proposal focus on the application of these technologies to study (A) how neuropeptides mediate experience dependent modulation of oxygen chemosensory circuits; and (B) how food availability and nutritional status modulate the same neural circuits. Aim (C) is an innovative engineering approach in which I will develop new microfluidic technologies that allow the simultaneous recording of oxygen evoked behaviors and neural activity. This will be beneficial for aims A and B and will pave way for new future research directions.

An animal’s decision on how to respond to the environment is based not only on the sensory information available, but further depends on internal factors such as stress, sleep / wakefulness, hunger / satiety and experience. Neurotransmitters and neuropeptides in the brain modulate neural circuits accordingly so that appropriate behaviors are generated. Aberrant neuromodulation is implicated in diseases such as insomnia, obesity or anorexia. Given the complexity of most neural systems studied, we lack good models of how neuromodulators systemically affect the activities of neural networks.
To overcome this problem, I propose to study neural circuits in the nematode C. elegans, which is a genetically tractable model organism with a simple and anatomically defined nervous system. I will focus on the neural circuits involved in oxygen chemosensory behaviors. Worms can smell oxygen and they use this information to navigate through heterogeneous environments. This enables them to find food and to engage in social interactions. Oxygen chemosensory behaviors are highly modulated by experience and nutritional status, but the underlying mechanisms are not understood.
I established behavioral assays that allow studying the modulation of oxygen behaviors in a rigorously quantifiable manner. I also acquired expertise in micro-fabrication technologies and developed imaging devices to measure the activity of neurons in live animals. The first two aims of this proposal focus on the application of these technologies to study (A) how neuropeptides mediate experience dependent modulation of oxygen chemosensory circuits; and (B) how food availability and nutritional status modulate the same neural circuits. Aim (C) is an innovative engineering approach in which I will develop new microfluidic technologies that allow the simultaneous recording of oxygen evoked behaviors and neural activity. This will be beneficial for aims A and B and will pave way for new future research directions.

Summary"The stereoselective preparation of enantioenriched organic compounds of high structural complexity and synthetic value, in an economically viable and expeditious manner, is one of the most important goals in contemporary Organic Synthesis. In this proposal, I present a unified and conceptually novel approach for the conversion of flat, aromatic heterocycles into highly valuable compounds for a variety of applications. This approach hinges upon a synergistic combination of the dramatic power of organic photochemical transformations combined with the exceedingly high selectivity and atom-economy of efficient catalytic processes. Indeed, the use of probably the cheapest reagent (light) combined with a catalytic transformation ensures near perfect atom-economy in this journey from flat and inexpensive substructures to chiral added-value products. Conceptually, the photochemical operation is envisaged as a energy-loading step whereas the catalytic transformation functions as an energy-release where asymmetric information is inscribed into the products.
The chemistry proposed herein will open up new vistas in enantioselective synthesis. Furthermore, groundbreaking and unprecedented methodology in the field of catalytic allylic alkylation is proposed that significantly expands (and goes beyond) the currently accepted “dogmas” for these textbook reactions. These include (but are not limited to) systematic violations of well-established rules “by design”, new contexts for application, new activation modes and innovative leaving groups. Finally, the comprehensive body of synthetic technology presented will be applied to pressing target-oriented problems in Organic Synthesis. It shall result in a landmark, highly efficient total synthesis of Tamiflu, as well as in application to an environmentally important target (Fomannosin), allowing the easy production of analogues for biological testing."

"The stereoselective preparation of enantioenriched organic compounds of high structural complexity and synthetic value, in an economically viable and expeditious manner, is one of the most important goals in contemporary Organic Synthesis. In this proposal, I present a unified and conceptually novel approach for the conversion of flat, aromatic heterocycles into highly valuable compounds for a variety of applications. This approach hinges upon a synergistic combination of the dramatic power of organic photochemical transformations combined with the exceedingly high selectivity and atom-economy of efficient catalytic processes. Indeed, the use of probably the cheapest reagent (light) combined with a catalytic transformation ensures near perfect atom-economy in this journey from flat and inexpensive substructures to chiral added-value products. Conceptually, the photochemical operation is envisaged as a energy-loading step whereas the catalytic transformation functions as an energy-release where asymmetric information is inscribed into the products.
The chemistry proposed herein will open up new vistas in enantioselective synthesis. Furthermore, groundbreaking and unprecedented methodology in the field of catalytic allylic alkylation is proposed that significantly expands (and goes beyond) the currently accepted “dogmas” for these textbook reactions. These include (but are not limited to) systematic violations of well-established rules “by design”, new contexts for application, new activation modes and innovative leaving groups. Finally, the comprehensive body of synthetic technology presented will be applied to pressing target-oriented problems in Organic Synthesis. It shall result in a landmark, highly efficient total synthesis of Tamiflu, as well as in application to an environmentally important target (Fomannosin), allowing the easy production of analogues for biological testing."

Max ERC Funding

1 487 000 €

Duration

Start date: 2012-01-01, End date: 2016-12-31

Project acronymHAPLOID

Project“Yeast” genetics in mammalian cells to identify fundamental mechanisms of physiology and pathophysiology

Researcher (PI)Josef Penninger

Host Institution (HI)INSTITUT FUER MOLEKULARE BIOTECHNOLOGIE GMBH

Call DetailsAdvanced Grant (AdG), LS4, ERC-2013-ADG

Summary"Some organisms such as yeast or social insects are haploid, i.e. they carry a single set of chromosomes. Organisms with a single copy of their genome provide a basis for genetic analyses where any recessive mutation of essential genes will show a clear phenotype due to the absence of a second gene copy. Recessive genetic screens have markedly contributed to our understanding of normal development, basic physiology, and disease. However, all somatic mammalian cells carry two copies of chromosomes (diploidy) that obscure mutational screens. Although deemed impossible, we were able to develop generate mammalian haploid embryonic stem cells, thereby breaking a paradigm of biology.
Our novel stem opens the possibility of combining the power of a haploid genome with pluripotency of embryonic stem cells to uncover fundamental biological processes in defined cell types at a genomic scale. The following projects are proposed:
1. Towards“yeast” genetics in mammalian stem cells. Development of optimized technologies for rapid, genome-wide screens via repairable mutagenesis. Mutational bar-coding to introduce quantitative genomics to mammalian biology.
2. Forward genetic screens to uncover essential stem cell genes, identify novel stemness factors, develop improved systems for iPS cell derivation, and to perform synthetic lethal screens for anti-cancer drugs.
3. Reverse genetics using to identify and validate genes involved in cardiovascular physiology, brown and white fat cell development, and pain sensing.
4. Hit validation – exemplified by resistance to the bioweapon ricin.
Haploid embryonic stem cells carry the promise to revolutionize functional genetics and allow rapid, near whole genome-wide mutational forward genetics analysis and reverse genetics in defined cell types. Our systems will be made available to all researchers and the knowledge gained from our studies should fundamentally impact on the basic understanding of physiology and disease pathogenesis."

"Some organisms such as yeast or social insects are haploid, i.e. they carry a single set of chromosomes. Organisms with a single copy of their genome provide a basis for genetic analyses where any recessive mutation of essential genes will show a clear phenotype due to the absence of a second gene copy. Recessive genetic screens have markedly contributed to our understanding of normal development, basic physiology, and disease. However, all somatic mammalian cells carry two copies of chromosomes (diploidy) that obscure mutational screens. Although deemed impossible, we were able to develop generate mammalian haploid embryonic stem cells, thereby breaking a paradigm of biology.
Our novel stem opens the possibility of combining the power of a haploid genome with pluripotency of embryonic stem cells to uncover fundamental biological processes in defined cell types at a genomic scale. The following projects are proposed:
1. Towards“yeast” genetics in mammalian stem cells. Development of optimized technologies for rapid, genome-wide screens via repairable mutagenesis. Mutational bar-coding to introduce quantitative genomics to mammalian biology.
2. Forward genetic screens to uncover essential stem cell genes, identify novel stemness factors, develop improved systems for iPS cell derivation, and to perform synthetic lethal screens for anti-cancer drugs.
3. Reverse genetics using to identify and validate genes involved in cardiovascular physiology, brown and white fat cell development, and pain sensing.
4. Hit validation – exemplified by resistance to the bioweapon ricin.
Haploid embryonic stem cells carry the promise to revolutionize functional genetics and allow rapid, near whole genome-wide mutational forward genetics analysis and reverse genetics in defined cell types. Our systems will be made available to all researchers and the knowledge gained from our studies should fundamentally impact on the basic understanding of physiology and disease pathogenesis."

Max ERC Funding

2 499 951 €

Duration

Start date: 2014-02-01, End date: 2019-01-31

Project acronymHIPECMEM

ProjectMemory-Related Information Processing in Neuronal Circuits of the Hippocampus and Entorhinal Cortex

Researcher (PI)Jozsef Csicsvari

Host Institution (HI)INSTITUTE OF SCIENCE AND TECHNOLOGY AUSTRIA

Call DetailsStarting Grant (StG), LS5, ERC-2011-StG_20101109

SummaryThis proposal will elucidate the circuit mechanism that underlies the spatial memory-related information processing in the interconnected brain areas of the hippocampus and entorhinal cortex (EC). Both of these areas are implicated in spatial memory and encode spatial information in neuronal activity patterns. The mechanisms underlying the emergence and coordination of spatial memory-related activity in these regions are needed to understand how these circuits process mnemonic information. Accordingly, here we aim at elucidating the representation of spatial memory by investigating these mechanisms at the circuit and synaptic levels of organisation. The first objective of this proposal is to characterise oscillatory synchronisation in hippocampo-EC circuits at different stages of memory processing. We hypothesise that network oscillations facilitate circuit interactions during memory processing. Therefore, using optogenetic techniques to disrupt oscillations, we aim at identifying critical periods during mnemonic processing when synchronisation is needed. Secondly, we intend to reveal how mnemonic information is encoded and exchanged between different areas of the hippocampo-EC system. We will test whether spatial memory-associated firing of dorsal hippocampal cells could be triggered by EC and/or ventral hippocampal cells that encode similar mnemonic features. In addition, this project will explore the role of temporal coding in the representation and consolidation of spatial memory traces. The third objective will investigate synaptic changes between connected CA3-CA3 and CA3-CA1 cell pairs during spatial learning. We will use cross-correlation analysis and electrical microstimulation to examine the rules that govern changes in synaptic efficacy by observing the probability of spike transmission.
Overall, the proposal provides a comprehensive approach to understanding how hippocampo-EC circuits organise and store information during mnemonic processes.

This proposal will elucidate the circuit mechanism that underlies the spatial memory-related information processing in the interconnected brain areas of the hippocampus and entorhinal cortex (EC). Both of these areas are implicated in spatial memory and encode spatial information in neuronal activity patterns. The mechanisms underlying the emergence and coordination of spatial memory-related activity in these regions are needed to understand how these circuits process mnemonic information. Accordingly, here we aim at elucidating the representation of spatial memory by investigating these mechanisms at the circuit and synaptic levels of organisation. The first objective of this proposal is to characterise oscillatory synchronisation in hippocampo-EC circuits at different stages of memory processing. We hypothesise that network oscillations facilitate circuit interactions during memory processing. Therefore, using optogenetic techniques to disrupt oscillations, we aim at identifying critical periods during mnemonic processing when synchronisation is needed. Secondly, we intend to reveal how mnemonic information is encoded and exchanged between different areas of the hippocampo-EC system. We will test whether spatial memory-associated firing of dorsal hippocampal cells could be triggered by EC and/or ventral hippocampal cells that encode similar mnemonic features. In addition, this project will explore the role of temporal coding in the representation and consolidation of spatial memory traces. The third objective will investigate synaptic changes between connected CA3-CA3 and CA3-CA1 cell pairs during spatial learning. We will use cross-correlation analysis and electrical microstimulation to examine the rules that govern changes in synaptic efficacy by observing the probability of spike transmission.
Overall, the proposal provides a comprehensive approach to understanding how hippocampo-EC circuits organise and store information during mnemonic processes.

Max ERC Funding

1 441 119 €

Duration

Start date: 2011-11-01, End date: 2016-10-31

Project acronymLipoCheX

ProjectThe Role of Lipolysis in the Pathogenesis of
Cancer-associated Cachexia

Researcher (PI)Rudolf Zechner

Host Institution (HI)UNIVERSITAET GRAZ

Call DetailsAdvanced Grant (AdG), LS4, ERC-2013-ADG

SummaryCachexia is a complex syndrome characterized by massive loss of body weight due to uncontrolled loss of adipose tissue and skeletal muscle. The wasting occurs during late stages of many unrelated chronic diseases and frequently leads to the death of affected individuals. Cachexia is most common in cancer, where an estimated 25% of patients die from cancer-associated cachexia (CAC) rather than from the cancer. Despite the tremendous impact of CAC on morbidity and mortality, the underlying molecular mechanisms are poorly understood.
Recently, we demonstrated that lipase-catalyzed triacylglycerol (TG) catabolism is required for the pathogenesis of CAC. Mice lacking adipose triglyceride lipase, the rate-limiting enzyme for TG hydrolysis (lipolysis), were completely protected from loss of both adipose tissue and muscle in two forms of cancer. This implies an essential role of the lipolytic process in the pathogenesis of CAC. Here we propose to elucidate the causal role of lipases and their coregulators in CAC development. We will determine mechanisms involved and pursue novel treatment strategies.
Our objectives are to:
- Investigate how different cancers in mice regulate tissue-specific lipolysis;
- Elucidate the functional role of lipases and their coregulators in the pathogenesis of CAC;
- Assess whether pharmacological inhibition of specific lipases prevents or delays CAC;
- Study the effects of cancer-induced lipolysis on energy dissipating pathways and epigenetic control.
The project enters a largely unexplored field: the role of lipid metabolism in the pathogenesis of CAC. The work will heavily rely on the characterization of induced mutant mouse models with CAC and require extensive collaboration with experts in pathology and large-scale systems analytics. The results are expected to yield new mechanisms of disease development and provide novel therapeutic targets to prevent the devastating and prevalent consequences of CAC.

Cachexia is a complex syndrome characterized by massive loss of body weight due to uncontrolled loss of adipose tissue and skeletal muscle. The wasting occurs during late stages of many unrelated chronic diseases and frequently leads to the death of affected individuals. Cachexia is most common in cancer, where an estimated 25% of patients die from cancer-associated cachexia (CAC) rather than from the cancer. Despite the tremendous impact of CAC on morbidity and mortality, the underlying molecular mechanisms are poorly understood.
Recently, we demonstrated that lipase-catalyzed triacylglycerol (TG) catabolism is required for the pathogenesis of CAC. Mice lacking adipose triglyceride lipase, the rate-limiting enzyme for TG hydrolysis (lipolysis), were completely protected from loss of both adipose tissue and muscle in two forms of cancer. This implies an essential role of the lipolytic process in the pathogenesis of CAC. Here we propose to elucidate the causal role of lipases and their coregulators in CAC development. We will determine mechanisms involved and pursue novel treatment strategies.
Our objectives are to:
- Investigate how different cancers in mice regulate tissue-specific lipolysis;
- Elucidate the functional role of lipases and their coregulators in the pathogenesis of CAC;
- Assess whether pharmacological inhibition of specific lipases prevents or delays CAC;
- Study the effects of cancer-induced lipolysis on energy dissipating pathways and epigenetic control.
The project enters a largely unexplored field: the role of lipid metabolism in the pathogenesis of CAC. The work will heavily rely on the characterization of induced mutant mouse models with CAC and require extensive collaboration with experts in pathology and large-scale systems analytics. The results are expected to yield new mechanisms of disease development and provide novel therapeutic targets to prevent the devastating and prevalent consequences of CAC.

Max ERC Funding

2 499 446 €

Duration

Start date: 2014-04-01, End date: 2019-03-31

Project acronymLUbi

ProjectRegulation and function of linear ubiquitination by HOIP

Researcher (PI)Fumiyo Ikeda

Host Institution (HI)INSTITUT FUER MOLEKULARE BIOTECHNOLOGIE GMBH

Call DetailsConsolidator Grant (CoG), LS1, ERC-2013-CoG

SummaryUbiquitin (Ub) is a small protein modifier, regulating diverse biological functions such as signalling, DNA repair and proteasomal degradation. Ub can form polymers via 7 Lys residues of Ub itself. Recently, we have discovered that an E3 ligase complex, Linear Ubiquitin chain Assembly Complex (LUBAC) generates a novel type of Ub polymer linked via Met-1, ‘linear Ub chain’ and regulates NF-kB signalling in mice. Because linear Ub is unique and the study of it is still in infancy, the only E3 ligase known is LUBAC, comprising a catalytic protein HOIP, and two regulatory subunits SHARPIN and HOIL-1L. We have shown that SHARPIN deficiency leads inflammation in mice. A mutation in HOIL-1L gene of human was shown to lead immunodeficiency. Yet, the regulatory mechanisms of HOIP catalytic activity and the biological implications remain poorly understood. Here, we aim to
- Elucidate the roles of HOIP in Drosophila
- Elucidate the roles of ubiquitination and ligase activities of mammalian HOIP in vivo
- Identify novel substrates of human HOIP and clarification of their roles
We recently identified an orthologue of HOIP in Drosophila, yet its genome does not encode SHARPIN or HOIL-1L. We aim to elucidate how dmHOIP mediates linear ubiquitination in the absence of regulatory subunits and the roles of HOIP in the NF-kB signalling by genetically deleting HOIP in Drosophila. We further aim to elucidate the role of HOIP E3 ligase activity and ubiquitination in inflammation by generating the conditional knockin mice of HOIP mutants. Moreover, we will develop a protein chip assay to identify new substrates of HOIP and determine how they contribute to the biological functions.
Since Ub plays such a wide variety of pathological functions including cancer, inflammation and neuronal diseases, I believe the expected results not only will lead to a better understanding of functional role of HOIP but will also identify novel aspects of linear ubiquitination in human diseases.

Ubiquitin (Ub) is a small protein modifier, regulating diverse biological functions such as signalling, DNA repair and proteasomal degradation. Ub can form polymers via 7 Lys residues of Ub itself. Recently, we have discovered that an E3 ligase complex, Linear Ubiquitin chain Assembly Complex (LUBAC) generates a novel type of Ub polymer linked via Met-1, ‘linear Ub chain’ and regulates NF-kB signalling in mice. Because linear Ub is unique and the study of it is still in infancy, the only E3 ligase known is LUBAC, comprising a catalytic protein HOIP, and two regulatory subunits SHARPIN and HOIL-1L. We have shown that SHARPIN deficiency leads inflammation in mice. A mutation in HOIL-1L gene of human was shown to lead immunodeficiency. Yet, the regulatory mechanisms of HOIP catalytic activity and the biological implications remain poorly understood. Here, we aim to
- Elucidate the roles of HOIP in Drosophila
- Elucidate the roles of ubiquitination and ligase activities of mammalian HOIP in vivo
- Identify novel substrates of human HOIP and clarification of their roles
We recently identified an orthologue of HOIP in Drosophila, yet its genome does not encode SHARPIN or HOIL-1L. We aim to elucidate how dmHOIP mediates linear ubiquitination in the absence of regulatory subunits and the roles of HOIP in the NF-kB signalling by genetically deleting HOIP in Drosophila. We further aim to elucidate the role of HOIP E3 ligase activity and ubiquitination in inflammation by generating the conditional knockin mice of HOIP mutants. Moreover, we will develop a protein chip assay to identify new substrates of HOIP and determine how they contribute to the biological functions.
Since Ub plays such a wide variety of pathological functions including cancer, inflammation and neuronal diseases, I believe the expected results not only will lead to a better understanding of functional role of HOIP but will also identify novel aspects of linear ubiquitination in human diseases.

Summary"Numerous scientific studies have established that the lunar cycle synchronizes reproductive behaviour and sexual maturation of animals as diverse as corals, polychaetes and fishes. Classical and recent work shows that in animals such as the annelid Platynereis dumerilii, dim nocturnal light serves as entrainment cue for an endogenous oscillator – a circalunar clock – that orchestrates reproductive and behavioral cycles. As circalunar clocks run with a (semi-)monthly period, they represent a fundamental biological phenomenon clearly distinct from the widely studied, solar light-entrained circadian (24h) clocks. Despite the vital importance of circalunar clocks, very little is known about the underlying molecular processes and responsible neuron types. This knowledge gap reflects the fact that until now, no suitable model system has been available to study circalunar clocks on the molecular and cellular level.
This proposal takes full advantage of the recent establishment of substantial molecular resources and critical techniques for functional analyses in Platynereis, as well as our pioneering work on the first circalunar clock-regulated genes and the identification of four molecular candidates for the nocturnal light receptor.
This now allows us to tackle two fundamental objectives:
First, we aim to discover the molecular and cellular nature of the lunar light sensor(s) and their interplay with solar light photoreceptors.
Second, we aim to characterize circalunar oscillatory genes and their associated neuron types that
will pave the way to unravel the molecular and cellular nature of the circalunar oscillator.
This work will provide new mechanistic insight into an unexplored biological mystery- circalunar clocks and their regulation by light. It also offers new conceptual advance into how animals accomplish the separation of diurnal versus nocturnal light information for the synchronization of reproductive behaviour, a challenge common in the natural environment."

"Numerous scientific studies have established that the lunar cycle synchronizes reproductive behaviour and sexual maturation of animals as diverse as corals, polychaetes and fishes. Classical and recent work shows that in animals such as the annelid Platynereis dumerilii, dim nocturnal light serves as entrainment cue for an endogenous oscillator – a circalunar clock – that orchestrates reproductive and behavioral cycles. As circalunar clocks run with a (semi-)monthly period, they represent a fundamental biological phenomenon clearly distinct from the widely studied, solar light-entrained circadian (24h) clocks. Despite the vital importance of circalunar clocks, very little is known about the underlying molecular processes and responsible neuron types. This knowledge gap reflects the fact that until now, no suitable model system has been available to study circalunar clocks on the molecular and cellular level.
This proposal takes full advantage of the recent establishment of substantial molecular resources and critical techniques for functional analyses in Platynereis, as well as our pioneering work on the first circalunar clock-regulated genes and the identification of four molecular candidates for the nocturnal light receptor.
This now allows us to tackle two fundamental objectives:
First, we aim to discover the molecular and cellular nature of the lunar light sensor(s) and their interplay with solar light photoreceptors.
Second, we aim to characterize circalunar oscillatory genes and their associated neuron types that
will pave the way to unravel the molecular and cellular nature of the circalunar oscillator.
This work will provide new mechanistic insight into an unexplored biological mystery- circalunar clocks and their regulation by light. It also offers new conceptual advance into how animals accomplish the separation of diurnal versus nocturnal light information for the synchronization of reproductive behaviour, a challenge common in the natural environment."

Max ERC Funding

1 500 000 €

Duration

Start date: 2014-02-01, End date: 2019-01-31

Project acronymmiRLIFE

ProjectMolecular Characterization of the microRNA Life-Cycle

Researcher (PI)Stefan Ludwig Ameres

Host Institution (HI)INSTITUT FUER MOLEKULARE BIOTECHNOLOGIE GMBH

Call DetailsStarting Grant (StG), LS1, ERC-2013-StG

SummarySmall silencing RNAs regulate gene expression in nearly all eukaryotes and have enormous biotechnological and therapeutic potential. MicroRNAs belong to the larges family of trans-acting gene regulatory molecules in multicellular organisms. In flies and mammals, they control more than half of the protein-coding transcriptome, and act as key regulators of organismal development, physiology, and disease.
Here, we propose to study the molecular mechanisms that regulate microRNA homeostasis. We aim to understand how distinct small RNA profiles are established and maintained to coordinate the expression of more than half of all protein coding genes in flies and mammals. Our studies will provide insight into the processes that regulate the function of miRNAs, determine possible causes for aberrant miRNA levels, that have been associated with human diseases, and provide guidelines how to efficiently inhibit miRNA function for analytical and therapeutic purposes.
We aim to identify and characterize the molecular determinants of microRNA stability, to dissect the pathways that promote the sequence-specific degradation of microRNAs, and to understand the biological consequences and therapeutic potential of small RNA decay. We will develop novel tools to obtain a view on the intracellular dynamics of RNA silencing pathways, in order to determine the molecular features associated with small RNA biogenesis and decay.
Because of its genetic and biochemical tools, we will use Drosophila melanogaster as a model organism. We will employ a combination of bioinformatics, cell-free biochemical experiments, cell culture methods, and in vivo genetics. What we learn in flies we will test in vitro in mammalian cell extracts, in cultured human cell lines and in vivo in mice to identify where these processes are conserved and where they diverge.
Overall, our goal is to determine fundamental biological mechanisms of RNA silencing, a phenomenon with enormous biological and biomedical impact.

Small silencing RNAs regulate gene expression in nearly all eukaryotes and have enormous biotechnological and therapeutic potential. MicroRNAs belong to the larges family of trans-acting gene regulatory molecules in multicellular organisms. In flies and mammals, they control more than half of the protein-coding transcriptome, and act as key regulators of organismal development, physiology, and disease.
Here, we propose to study the molecular mechanisms that regulate microRNA homeostasis. We aim to understand how distinct small RNA profiles are established and maintained to coordinate the expression of more than half of all protein coding genes in flies and mammals. Our studies will provide insight into the processes that regulate the function of miRNAs, determine possible causes for aberrant miRNA levels, that have been associated with human diseases, and provide guidelines how to efficiently inhibit miRNA function for analytical and therapeutic purposes.
We aim to identify and characterize the molecular determinants of microRNA stability, to dissect the pathways that promote the sequence-specific degradation of microRNAs, and to understand the biological consequences and therapeutic potential of small RNA decay. We will develop novel tools to obtain a view on the intracellular dynamics of RNA silencing pathways, in order to determine the molecular features associated with small RNA biogenesis and decay.
Because of its genetic and biochemical tools, we will use Drosophila melanogaster as a model organism. We will employ a combination of bioinformatics, cell-free biochemical experiments, cell culture methods, and in vivo genetics. What we learn in flies we will test in vitro in mammalian cell extracts, in cultured human cell lines and in vivo in mice to identify where these processes are conserved and where they diverge.
Overall, our goal is to determine fundamental biological mechanisms of RNA silencing, a phenomenon with enormous biological and biomedical impact.